We described rickets in children in our inner city area (over 80% minorities) with either low or normal circulating 25-hydroxyvitamin D (25-OHD) levels, and have found that vitamin D deficiency (circulating 25-OHD <20 ng/ml) occurs in 13% of over 750 6 mo. to 3 yr-old healthy children sampled in this setting. Moreover, our recent analysis of vitamin D status in these children reveals that vitamin D binding protein (DBP) genotype is a significant predictor of 25-OHD and 1,25 dihydroxyvitamin D [1,25 (OH)2D] levels. When examining the two most prevalent minorities in this group, Hispanic and African-American, significant interactions between ethnicity and DBP genotype are evident. We therefore suspect that vitamin D requirements may vary according to DBP genotype, and amongst minority ethnic groups. Moreover DBP genotype may be important for both interpretation of vitamin D levels and the physiologic response to vitamin D supplementation. Likewise, a DBP effect on susceptibility to vitamin D intoxication may be apparent. Therefore our study will examine physiologic and biochemical responses to vitamin D supplementation in a cohort of primarily minority children, most susceptible to overt vitamin D deficiency, and stratified by DBP genotypes. To examine responses we will focus on both classical and novel non-classical effects of vitamin D. Hypothesis 1. Response to vitamin D supplementation may differ among children of varying DBP genotypes. If so, recommendations for optimal vitamin D supplementation may need to consider DBP genotype. We will explore this possibility by determining the biochemical response to different levels of vitamin D supplementation (400 and 1000 units daily). We will enroll children of 3 genotypes associated with the most extreme differences in circulating levels of 25-OHD and 1,25(OH)2D from our extensively phenotyped cohort of over 750 subjects (and recruit additional subjects as needed) to perform a randomized trial of vitamin D supplementation at these 2 dosing levels, stratified by the 3 DBP genotype groups. This specific aim will aid in establishing appropriate dosing for infants and toddlers in the demographic most closely associated with reports of vitamin D deficiency by assessing the achievement of target levels of 25-OHD in the selected genotypes at the two dosing levels. Circulating 1,25 (OH)2D will also be examined as a secondary endpoint. Hypothesis 2. Although our preliminary data indicate that vitamin D metabolite levels differ across genotypes, it is not clear if the functional consequences of vitamin D actions are determined by DBP genotype. Therefore, we will examine the effect of vitamin D supplementation performed in Specific Aim 2 on: a) classical endocrine effects of vitamin D on the bone and mineral system, with outcome measures to include: circulating levels of PTH, alkaline phosphatase activity, osteocalcin, P1NP, and FGF23 as markers of a vitamin D effect on the skeleton and bone mineral density by ultrasound;and b) non- classical effects of vitamin D including changes in BMI, incidence of infection, blood pressure, insulin sensitivity (using measures derived from fasting insulin and cortisol levels) and circulating levels of aldosterone and renin. Finally, we will examine effects of vitamin D supplementation on the immune response of peripheral blood mononuclear cells. By assessing expression of the vitamin D-dependent antimicrobial peptide, cathelicidin before and after vitamin D supplementation, this study will provide data for establishing physiologically sound recommendations for safe, effective vitamin D supplementation in children. We will explore the possibility that DBP genotype may affect the response to supplementation. The data will directly impact the population from which the subjects are recruited, those children between 6 months and 6 years of age, in urban environments and at greatest risk for developing vitamin D deficiency. Moreover, the data are likely to serve the larger pediatric population as well. We expect that in 3-4 years the data could be disseminated for directing development of policy, and to serve as guidelines for practice. PUBLIC HEALTH RELEVANCE: This study will provide data for establishing physiologically sound recommendations for safe, effective vitamin D supplementation in children. We will explore the possibility that DBP genotype may affect the response to supplementation. The data will directly impact the population from which the subjects are recruited, those children between 6 months and 6 years of age, in urban environments and at greatest risk for developing vitamin D deficiency. Moreover, the data are likely to serve the larger pediatric population as well.